流感病毒包膜蛋白M2免疫防护能力和防护机制研究
发布时间：2019-03-31 13:57
【摘要】： 流感的流行给人类健康带来很大的危害,这就迫切要求开发出具有广谱防护效果的流感疫苗。流感病毒包膜蛋白M2因其高度保守性而成为疫苗研制的重要靶蛋白。在本论文中,我们重点研究了流感病毒M2蛋白胞外区(M2e)的免疫防护能力和防护机制,并取得了以下创新性研究成果。 我们发现流感M2e具有很强的免疫原性,合成的M2e多肽无需载体蛋白就能诱导产生高滴度的特异性抗体,并且在攻毒实验中能保护小鼠免于致死剂量流感病毒的攻击。此外,研究发现M2e多肽能诱导产生特异性的T细胞反应,从而证明了在流感病毒M2e上存在Th细胞表位。这些研究成果部分阐明了流感病毒M2蛋白的免疫防护机制。 研究发现利用定点突变的方法将M2e多肽上的半胱氨酸残基替换成丝氨酸后不会影响M2e多肽的抗原性,但会显著降低其免疫原性,丧失诱导产生特异性免疫应答的能力。此外,我们还发现M2e多肽可以通过半胱氨酸残基形成的链间二硫键交联形成多聚体。这表明M2e的半胱氨酸残基对维持其免疫原性起到关键作用。 为了探讨M2蛋白诱导的免疫应答与病毒防护效果之间的关系,我们利用添加CpG-ODN佐剂的方法来影响M2e多肽诱导产生的免疫应答。研究发现在M2e多肽中加入CpG-ODN能显著提高其诱导产生的抗体水平和T细胞反应,并且促进免疫反应向Th1型发展。但是免疫应答的增强反而降低了高剂量病毒攻击时的防护效果。 我们将M2e多肽与传统的裂解疫苗联合免疫,以增强裂解疫苗对不同亚型毒株的交叉防护能力。研究发现M2e多肽与裂解疫苗的联合免疫不会影响血凝抑制抗体的产生,但是佐剂的使用对联合免疫诱导的M2e特异性免疫应答有重要影响。在铝佐剂中,联合免疫能诱导产生M2e特异性的免疫应答,而在MF59佐剂中却无此效果。此外,疫苗诱导的交叉防护效果也受所用佐剂的影响。在铝佐剂中,加入M2e多肽能提高裂解疫苗的交叉防护效果,而在MF59佐剂中反而降低其交叉防护能力。这些研究结果提示M2实验疫苗的防护效果也与佐剂的配伍相关。
[Abstract]:Influenza epidemic has brought great harm to human health, so it is urgent to develop influenza vaccine with broad-spectrum protective effect. Influenza virus envelope protein M2 has become an important target protein for vaccine development because of its high conservatism. In this paper, we focus on the immune protection ability and protection mechanism of influenza virus M2 protein extracellular domain (M2e), and obtain the following innovative research results. We found that influenza M2e has strong immunogenicity. The synthesized M2e peptide can induce high titer of specific antibody without vector protein, and can protect mice from lethal dose of influenza virus in the challenge experiment. In addition, it was found that M2e peptide could induce specific T cell response, which confirmed the existence of Th cell epitopes on influenza virus M2e. These results partly clarify the immune protection mechanism of influenza virus M2 protein. It was found that the substitution of cysteine residue from M _ 2e peptide with serine by site-directed mutation did not affect the antigenicity of M _ 2e peptide, but significantly decreased its immunogenicity and lost the ability to induce specific immune response. In addition, we also found that M _ 2e peptide can cross-link with disulfide bonds formed by cysteine residues to form polymers. This suggests that the cysteine residues of M2e play a key role in maintaining its immunogenicity. In order to investigate the relationship between the immune response induced by M2 protein and the protective effect of virus, we used the method of adding CpG-ODN adjuvant to influence the immune response induced by M2 peptide. It was found that the addition of CpG-ODN to M2e peptide could significantly increase the antibody level and T cell response induced by M2e peptide, and promote the development of immune response to Th1 type. However, the enhancement of immune response reduces the protective effect of high-dose virus attack. We combined the M2e peptide with the traditional lytic vaccine to enhance the cross-protection ability of the lytic vaccine against different subtypes of the vaccine. It was found that the co-immunization of M2e peptide and lytic vaccine did not affect the production of hemagglutination inhibition antibody, but the use of adjuvant had an important effect on the specific immune response of M2e induced by combined immunization. In aluminum adjuvant, co-immunization can induce M2e-specific immune response, but not in MF59 adjuvant. In addition, the cross-protective effect induced by the vaccine is also affected by the adjuvant used. In aluminum adjuvants, the addition of M _ 2e peptide could improve the cross-protection effect of lytic vaccine, while in MF59 adjuvant, the cross-protection ability of M2e peptide decreased. These results suggest that the protective effect of M2 experimental vaccine is also related to the compatibility of adjuvant.
【学位授予单位】：清华大学
【学位级别】：博士
【学位授予年份】：2008
【分类号】：R392
本文编号：2450979
[Abstract]:Influenza epidemic has brought great harm to human health, so it is urgent to develop influenza vaccine with broad-spectrum protective effect. Influenza virus envelope protein M2 has become an important target protein for vaccine development because of its high conservatism. In this paper, we focus on the immune protection ability and protection mechanism of influenza virus M2 protein extracellular domain (M2e), and obtain the following innovative research results. We found that influenza M2e has strong immunogenicity. The synthesized M2e peptide can induce high titer of specific antibody without vector protein, and can protect mice from lethal dose of influenza virus in the challenge experiment. In addition, it was found that M2e peptide could induce specific T cell response, which confirmed the existence of Th cell epitopes on influenza virus M2e. These results partly clarify the immune protection mechanism of influenza virus M2 protein. It was found that the substitution of cysteine residue from M _ 2e peptide with serine by site-directed mutation did not affect the antigenicity of M _ 2e peptide, but significantly decreased its immunogenicity and lost the ability to induce specific immune response. In addition, we also found that M _ 2e peptide can cross-link with disulfide bonds formed by cysteine residues to form polymers. This suggests that the cysteine residues of M2e play a key role in maintaining its immunogenicity. In order to investigate the relationship between the immune response induced by M2 protein and the protective effect of virus, we used the method of adding CpG-ODN adjuvant to influence the immune response induced by M2 peptide. It was found that the addition of CpG-ODN to M2e peptide could significantly increase the antibody level and T cell response induced by M2e peptide, and promote the development of immune response to Th1 type. However, the enhancement of immune response reduces the protective effect of high-dose virus attack. We combined the M2e peptide with the traditional lytic vaccine to enhance the cross-protection ability of the lytic vaccine against different subtypes of the vaccine. It was found that the co-immunization of M2e peptide and lytic vaccine did not affect the production of hemagglutination inhibition antibody, but the use of adjuvant had an important effect on the specific immune response of M2e induced by combined immunization. In aluminum adjuvant, co-immunization can induce M2e-specific immune response, but not in MF59 adjuvant. In addition, the cross-protective effect induced by the vaccine is also affected by the adjuvant used. In aluminum adjuvants, the addition of M _ 2e peptide could improve the cross-protection effect of lytic vaccine, while in MF59 adjuvant, the cross-protection ability of M2e peptide decreased. These results suggest that the protective effect of M2 experimental vaccine is also related to the compatibility of adjuvant.
【学位授予单位】：清华大学
【学位级别】：博士
【学位授予年份】：2008
【分类号】：R392
【引证文献】
相关博士学位论文 前1条
1 范克伟;Itk信号转导通路在A型流感病毒与T细胞相互作用中的功能研究[D];福建农林大学;2012年
，本文编号：2450979
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